Hunting for pigments in bacterial settlers of the Great Pacific Garbage Patch.

The Great Pacific Garbage Patch, a significant collection of plastic introduced by human activities, provides an ideal environment to study bacterial lifestyles on plastic substrates. We proposed that bacteria colonizing the floating plastic debris would develop strategies to deal with the ultraviolet-exposed substrate, such as the production of antioxidant pigments. We observed a variety of pigmentation in 67 strains that were directly cultivated from plastic pieces sampled from the Garbage Patch. The genomic analysis of four representative strains, each distinct in taxonomy, revealed multiple pathways for carotenoid production. These pathways include those that produce less common carotenoids and a cluster of photosynthetic genes. This cluster appears to originate from a potentially new species of the Rhodobacteraceae family. This represents the first report of an aerobic anoxygenic photoheterotrophic bacterium from plastic biofilms. Spectral analysis showed that the bacteria actively produce carotenoids, such as beta-carotene and beta-cryptoxanthin, and bacteriochlorophyll a. Furthermore, we discovered that the genetic ability to synthesize carotenoids is more common in plastic biofilms than in the surrounding water communities. Our findings suggest that plastic biofilms could be an overlooked source of bacteria-produced carotenoids, including rare forms. It also suggests that photoreactive molecules might play a crucial role in bacterial biofilm communities in surface water.

Blue valorization of lignin-derived monomers via reprogramming marine bacterium Roseovarius nubinhibens.

Biological valorization of lignin, the second most abundant biopolymer on Earth, is an indispensable sector to build a circular economy and net-zero future. However, lignin is recalcitrant to bioupcycling, demanding innovative solutions. We report here the biological valorization of lignin-derived aromatic carbon to value-added chemicals without requesting extra organic carbon and freshwater via reprogramming the marine Roseobacter clade bacterium Roseovarius nubinhibens. We discovered the unusual advantages of this strain for the oxidation of lignin monomers and implemented a CRISPR interference (CRISPRi) system with the lacI-Ptrc inducible module, nuclease-deactivated Cas9, and programmable gRNAs. This is the first CRISPR-based regulatory system in R. nubinhibens, enabling precise and efficient repression of genes of interest. By deploying the customized CRISPRi, we reprogrammed the carbon flux from a lignin monomer, 4-hydroxybenzoate, to achieve the maximum production of protocatechuate, a pharmaceutical compound with antibacterial, antioxidant, and anticancer properties, with minimal carbon to maintain cell growth and drive biocatalysis. As a result, we achieved a 4.89-fold increase in protocatechuate yield with a dual-targeting CRISPRi system, and the system was demonstrated with real seawater. Our work underscores the power of CRISPRi in exploiting novel microbial chassis and will accelerate the development of marine synthetic biology. Meanwhile, the introduction of a new-to-the-field lineage of marine bacteria unveils the potential of blue biotechnology leveraging resources from the ocean.IMPORTANCEOne often overlooked sector in carbon-conservative biotechnology is the water resource that sustains these enabling technologies. Similar to the "food-versus-fuel" debate, the competition of freshwater between human demands and bioproduction is another controversial issue, especially under global water scarcity. Here, we bring a new-to-the-field lineage of marine bacteria with unusual advantages to the stage of engineering biology for simultaneous carbon and water conservation. We report the valorization of lignin monomers to pharmaceutical compounds without requesting extra organic substrate (e.g., glucose) or freshwater by reprogramming the marine bacterium Roseovarius nubinhibens with a multiplex CRISPR interference system. Beyond the blue lignin valorization, we present a proof-of-principle of leveraging marine bacteria and engineering biology for a sustainable future.

Ligand cross-feeding resolves bacterial vitamin B12 auxotrophies.

Cobalamin (vitamin B12, herein referred to as B12) is an essential cofactor for most marine prokaryotes and eukaryotes1,2. Synthesized by a limited number of prokaryotes, its scarcity affects microbial interactions and community dynamics2-4. Here we show that two bacterial B12 auxotrophs can salvage different B12 building blocks and cooperate to synthesize B12. A Colwellia sp. synthesizes and releases the activated lower ligand α-ribazole, which is used by another B12 auxotroph, a Roseovarius sp., to produce the corrin ring and synthesize B12. Release of B12 by Roseovarius sp. happens only in co-culture with Colwellia sp. and only coincidently with the induction of a prophage encoded in Roseovarius sp. Subsequent growth of Colwellia sp. in these conditions may be due to the provision of B12 by lysed cells of Roseovarius sp. Further evidence is required to support a causative role for prophage induction in the release of B12. These complex microbial interactions of ligand cross-feeding and joint B12 biosynthesis seem to be widespread in marine pelagic ecosystems. In the western and northern tropical Atlantic Ocean, bacteria predicted to be capable of salvaging cobinamide and synthesizing only the activated lower ligand outnumber B12 producers. These findings add new players to our understanding of B12 supply to auxotrophic microorganisms in the ocean and possibly in other ecosystems.

Enantioselective transformation of phytoplankton-derived dihydroxypropanesulfonate by marine bacteria.

Chirality, a fundamental property of matter, is often overlooked in the studies of marine organic matter cycles. Dihydroxypropanesulfonate (DHPS), a globally abundant organosulfur compound, serves as an ecologically important currency for nutrient and energy transfer from phytoplankton to bacteria in the ocean. However, the chirality of DHPS in nature and its transformation remain unclear. Here, we developed a novel approach using chiral phosphorus-reagent labeling to separate DHPS enantiomers. Our findings demonstrated that at least one enantiomer of DHPS is present in marine diatoms and coccolithophores, and that both enantiomers are widespread in marine environments. A novel chiral-selective DHPS catabolic pathway was identified in marine Roseobacteraceae strains, where HpsO and HpsP dehydrogenases at the gateway to DHPS catabolism act specifically on R-DHPS and S-DHPS, respectively. R-DHPS is also a substrate for the dehydrogenase HpsN. All three dehydrogenases generate stable hydrogen bonds between the chirality-center hydroxyls of DHPS and highly conserved residues, and HpsP also form coordinate-covalent bonds between the chirality-center hydroxyls and Zn2+, which determines the mechanistic basis of strict stereoselectivity. We further illustrated the role of enzymatic promiscuity in the evolution of DHPS metabolism in Roseobacteraceae and SAR11. This study provides the first evidence of chirality's involvement in phytoplankton-bacteria metabolic currencies, opening a new avenue for understanding the ocean organosulfur cycle.

Rhodobacteraceae methanethiol oxidases catalyze methanethiol degradation to produce sulfane sulfur other than hydrogen sulfide.

Methanethiol (MT) is a sulfur-containing compound produced during dimethylsulfoniopropionate (DMSP) degradation by marine bacteria. The C-S bond of MT can be cleaved by methanethiol oxidases (MTOs) to release a sulfur atom. However, the cleaving process remains unclear, and the species of sulfur product is uncertain. It has long been assumed that MTOs produce hydrogen sulfide (H2S) from MT. Herein, we studied the MTOs in the Rhodobacteraceae family-whose members are important DMSP degraders ubiquitous in marine environments. We identified 57 MTOs from 1,904 Rhodobacteraceae genomes. These MTOs were grouped into two major clusters. Cluster 1 members share three conserved cysteine residues, while cluster 2 members contain one conserved cysteine residue. We examined the products of three representative MTOs both in vitro and in vivo. All of them produced sulfane sulfur other than H2S from MT. Their conserved cysteines are substrate-binding sites in which the MTO-S-S-CH3 complex is formed. This finding clarified the sulfur product of MTOs and enlightened the MTO-catalyzing process. Moreover, this study connected DMSP degradation with sulfane sulfur metabolism, filling a critical gap in the DMSP degradation pathway and representing new knowledge in the marine sulfur cycle field. IMPORTANCE: This study overthrows a long-time assumption that methanethiol oxidases (MTOs) cleave the C-S bond of methanethiol to produce both H2S and H2O2-the former is a strong reductant and the latter is a strong oxidant. From a chemistry viewpoint, this reaction is difficult to happen. Investigations on three representative MTOs indicated that sulfane sulfur (S0) was the direct product, and no H2O2 was produced. Finally, the products of MTOs were corrected to be S0 and H2O. This finding connected dimethylsulfoniopropionate (DMSP) degradation with sulfane sulfur metabolism, filling a critical gap in the DMSP degradation pathway and representing new knowledge in the marine sulfur cycle field.

Aminolipids elicit functional trade-offs between competitiveness and bacteriophage attachment in Ruegeria pomeroyi.

Lipids play a crucial role in maintaining cell integrity and homeostasis with the surrounding environment. Cosmopolitan marine roseobacter clade (MRC) and SAR11 clade bacteria are unique in that, in addition to glycerophospholipids, they also produce an array of amino acid-containing lipids that are conjugated with beta-hydroxy fatty acids through an amide bond. Two of these aminolipids, the ornithine aminolipid (OL) and the glutamine aminolipid (QL), are synthesized using the O-acetyltransferase OlsA. Here, we demonstrate that OL and QL are present in both the inner and outer membranes of the Gram-negative MRC bacterium Ruegeria pomeroyi DSS-3. In an olsA mutant, loss of these aminolipids is compensated by a concurrent increase in glycerophospholipids. The inability to produce aminolipids caused significant changes in the membrane proteome, with the membrane being less permeable and key nutrient transporters being downregulated while proteins involved in the membrane stress response were upregulated. Indeed, the import of 14C-labelled choline and dimethylsulfoniopropionate, as a proxy for the transport of key marine nutrients across membranes, was significantly impaired in the olsA mutant. Moreover, the olsA mutant was significantly less competitive than the wild type (WT) being unable to compete with the WT strain in co-culture. However, the olsA mutant unable to synthesize these aminolipids is less susceptible to phage attachment. Together, these data reveal a critical role for aminolipids in the ecophysiology of this important clade of marine bacteria and a trade-off between growth and avoidance of bacteriophage attachment.

Oxidative Stress Regulates a Pivotal Metabolic Switch in Dimethylsulfoniopropionate Degradation by the Marine Bacterium Ruegeria pomeroyi.

Dimethylsulfoniopropionate (DMSP) is an abundant organic compound in marine surface water and source of dimethyl sulfide (DMS), the largest natural sulfur source to the upper atmosphere. Marine bacteria either mineralize DMSP through the demethylation pathway or transform it to DMS through the cleavage pathway. Factors that regulate which pathway is utilized are not fully understood. In chemostat experiments, the marine Roseobacter Ruegeria pomeroyi DSS-3 was exposed to oxidative stress either during growth with H2O2 or by mutation of the gene encoding catalase. Oxidative stress reduced expression of the genes in the demethylation pathway and increased expression of those encoding the cleavage pathway. These results are contrary to the sulfur demand hypothesis, which theorizes that DMSP metabolism is driven by sulfur requirements of bacterial cells. Instead, we find strong evidence consistent with oxidative stress control over the switch in DMSP metabolism from demethylation to DMS production in an ecologically relevant marine bacterium. IMPORTANCE Dimethylsulfoniopropionate (DMSP) is the most abundant low-molecular-weight organic compound in marine surface water and source of dimethyl sulfide (DMS), a climatically active gas that connects the marine and terrestrial sulfur cycles. Marine bacteria are the major DMSP consumers, either generating DMS or consuming DMSP as a source of reduced carbon and sulfur. However, the factors regulating the DMSP catabolism in bacteria are not well understood. Marine bacteria are also exposed to oxidative stress. RNA sequencing (RNA-seq) experiments showed that oxidative stress induced in the laboratory reduced expression of the genes encoding the consumption of DMSP via the demethylation pathway and increased the expression of genes encoding DMS production via the cleavage pathway in the marine bacterium Ruegeria pomeroyi. These results support a model where DMS production in the ocean is regulated in part by oxidative stress.

Growth Substrate and Prophage Induction Collectively Influence Metabolite and Lipid Profiles in a Marine Bacterium.

Bacterial growth substrates influence a variety of biological functions, including the biosynthesis and regulation of lipid intermediates. The extent of this rewiring is not well understood nor has it been considered in the context of virally infected cells. Here, we used a one-host-two-temperate phage model system to probe the combined influence of growth substrate and phage infection on host carbon and lipid metabolism. Using untargeted metabolomics and lipidomics, we reported the detection of a suite of metabolites and lipid classes for two Sulfitobacter lysogens provided with three growth substrates of differing complexity and nutrient composition (yeast extract/tryptone [complex], glutamate and acetate). The growth medium led to dramatic differences in the detectable intracellular metabolites, with only 15% of 175 measured metabolites showing overlap across the three growth substrates. Between-strain differences were most evident in the cultures grown on acetate, followed by glutamate then complex medium. Lipid distribution profiles were also distinct between cultures grown on different substrates as well as between the two lysogens grown in the same medium. Five phospholipids, three aminolipid, and one class of unknown lipid-like features were identified. Most (≥94%) of these 75 lipids were quantifiable in all samples. Metabolite and lipid profiles were strongly determined by growth medium composition and modestly by strain type. Because fluctuations in availability and form of carbon substrates and nutrients, as well as virus pressure, are common features of natural systems, the influence of these intersecting factors will undoubtedly be imprinted in the metabolome and lipidome of resident bacteria. IMPORTANCE Community-level metabolomics approaches are increasingly used to characterize natural microbial populations. These approaches typically depend upon temporal snapshots from which the status and function of communities are often inferred. Such inferences are typically drawn from lab-based studies of select model organisms raised under limited growth conditions. To better interpret community-level data, the extent to which ecologically relevant bacteria demonstrate metabolic flexibility requires elucidation. Herein, we used an environmentally relevant model heterotrophic marine bacterium to assess the relationship between growth determinants and metabolome. We also aimed to assess the contribution of phage activity to the host metabolome. Striking differences in primary metabolite and lipid profiles appeared to be driven primarily by growth regime and, secondarily, by phage type. These findings demonstrated the malleable nature of metabolomes and lipidomes and lay the foundation for future studies that relate cellular composition with function in complex environmental microbial communities.

Dissolved organic phosphorus utilization by the marine bacterium Ruegeria pomeroyi DSS-3 reveals chain length-dependent polyphosphate degradation.

Dissolved organic phosphorus (DOP) is a critical nutritional resource for marine microbial communities. However, the relative bioavailability of different types of DOP, such as phosphomonoesters (P-O-C) and phosphoanhydrides (P-O-P), is poorly understood. Here we assess the utilization of these P sources by a representative bacterial copiotroph, Ruegeria pomeroyi DSS-3. All DOP sources supported equivalent growth by R. pomeroyi, and all DOP hydrolysis rates were upregulated under phosphorus depletion (-P). A long-chain polyphosphate (45polyP) showed the lowest hydrolysis rate of all DOP substrates tested, including tripolyphosphate (3polyP). Yet the upregulation of 45polyP hydrolysis under -P was greater than any other substrate analyzed. Proteomics revealed three common P acquisition enzymes potentially involved in polyphosphate utilization, including two alkaline phosphatases, PhoD and PhoX, and one 5'-nucleotidase (5'-NT). Results from DOP substrate competition experiments show that these enzymes likely have broad substrate specificities, including chain length-dependent reactivity toward polyphosphate. These results confirm that DOP, including polyP, are bioavailable nutritional P sources for R. pomeroyi, and possibly other marine heterotrophic bacteria. Furthermore, the chain-length dependent mechanisms, rates and regulation of polyP hydrolysis suggest that these processes may influence the composition of DOP and the overall recycling of nutrients within marine dissolved organic matter.

Role is in the eye of the beholder-the multiple functions of the antibacterial compound tropodithietic acid produced by marine Rhodobacteraceae.

Many microbial secondary metabolites have been studied for decades primarily because of their antimicrobial properties. However, several of these metabolites also possess nonantimicrobial functions, both influencing the physiology of the producer and their ecological neighbors. An example of a versatile bacterial secondary metabolite with multiple functions is the tropone derivative tropodithietic acid (TDA). TDA is a broad-spectrum antimicrobial compound produced by several members of the Rhodobacteraceae family, a major marine bacterial lineage, within the genera Phaeobacter, Tritonibacter, and Pseudovibrio. The production of TDA is governed by the mode of growth and influenced by the availability of nutrient sources. The antibacterial effect of TDA is caused by disruption of the proton motive force of target microorganisms and, potentially, by its iron-chelating properties. TDA also acts as a signaling molecule, affecting gene expression in other bacteria, and altering phenotypic traits such as motility, biofilm formation, and antibiotic production in the producer. In microbial communities, TDA-producing bacteria cause a reduction of the relative abundance of closely related species and some fast-growing heterotrophic bacteria. Here, we summarize the current understanding of the chemical ecology of TDA, including the environmental niches of TDA-producing bacteria, and the molecular mechanisms governing the function and regulation of TDA.

Structural Elucidation of Cryptic Algaecides in Marine Algal-Bacterial Symbioses by NMR Spectroscopy and MicroED.

Microbial secondary metabolite discovery is often conducted in pure monocultures. In a natural setting, however, where metabolites are constantly exchanged, biosynthetic precursors are likely provided by symbionts or hosts. In the current work, we report eight novel and architecturally unusual secondary metabolites synthesized by the bacterial symbiont Phaeobacter inhibens from precursors that, in a native context, would be provided by their algal hosts. Three of these were produced at low titres and their structures were determined de novo using the emerging microcrystal electron diffraction method. Some of the new metabolites exhibited potent algaecidal activity suggesting that the bacterial symbiont can convert algal precursors, tryptophan and sinapic acid, into complex cytotoxins. Our results have important implications for the parasitic phase of algal-bacterial symbiotic interactions.

Algal p-coumaric acid induces oxidative stress and siderophore biosynthesis in the bacterial symbiont Phaeobacter inhibens.

The marine alpha-proteobacterium Phaeobacter inhibens engages in intermittent symbioses with microalgae. The symbiosis is biphasic and concludes in a parasitic phase, during which the bacteria release algaecidal metabolites in response to algal p-coumaric acid (pCA). The cell-wide effects of pCA on P. inhibens remain unknown. Herein, we report a microarray-based transcriptomic study and find that genes related to the oxidative stress response and secondary metabolism are upregulated most, while those associated with energy production and motility are downregulated in the presence of pCA. Among genes upregulated is a previously unannotated biosynthetic gene cluster and, using a combination of gene deletions and metabolic profiling, we show that it gives rise to an unreported siderophore, roseobactin. The simultaneous production of algaecides and roseobactin in the parasitic phase allows the bacteria to take up any iron that is released from dying algal cells, thereby securing a limited micronutrient.

Acrylate protects a marine bacterium from grazing by a ciliate predator.

Cleavage of dimethylsulfoniopropionate (DMSP) can deter herbivores in DMSP-producing eukaryotic algae; however, it is unclear whether a parallel defence mechanism operates in marine bacteria. Here we demonstrate that the marine bacterium Puniceibacterium antarcticum SM1211, which does not use DMSP as a carbon source, has a membrane-associated DMSP lyase, DddL. At high concentrations of DMSP, DddL causes an accumulation of acrylate around cells through the degradation of DMSP, which protects against predation by the marine ciliate Uronema marinum. The presence of acrylate can alter the grazing preference of U. marinum to other bacteria in the community, thereby influencing community structure.

Genomic Evolution of the Marine Bacterium Phaeobacter inhibens during Biofilm Growth.

Phaeobacter inhibens 2.10 is an effective biofilm former on marine surfaces and has the ability to outcompete other microorganisms, possibly due to the production of the plasmid-encoded secondary metabolite tropodithietic acid (TDA). P. inhibens 2.10 biofilms produce phenotypic variants with reduced competitiveness compared to the wild type. In the present study, we used longitudinal, genome-wide deep sequencing to uncover the genetic foundation that contributes to the emergent phenotypic diversity in P. inhibens 2.10 biofilm dispersants. Our results show that phenotypic variation is not due to the loss of the plasmid that carries the genes for TDA synthesis but instead show that P. inhibens 2.10 biofilm populations become rapidly enriched in single nucleotide variations in genes involved in the synthesis of TDA. While variants in genes previously linked to other phenotypes, such as lipopolysaccharide production (i.e., rfbA) and cellular persistence (i.e., metG), also appear to be selected for during biofilm dispersal, the number and consistency of variations found for genes involved in TDA production suggest that this metabolite imposes a burden on P. inhibens 2.10 cells. Our results indicate a strong selection pressure for the loss of TDA in monospecies biofilm populations and provide insight into how competition (or a lack thereof) in biofilms might shape genome evolution in bacteria. IMPORTANCE Biofilm formation and dispersal are important survival strategies for environmental bacteria. During biofilm dispersal, cells often display stable and heritable variants from the parental biofilm. Phaeobacter inhibens is an effective colonizer of marine surfaces, in which a subpopulation of its biofilm dispersal cells displays a noncompetitive phenotype. This study aimed to elucidate the genetic basis of these phenotypic changes. Despite the progress made to date in characterizing the dispersal variants in P. inhibens, little is understood about the underlying genetic changes that result in the development of the specific variants. Here, P. inhibens phenotypic variation was linked to single nucleotide polymorphisms (SNPs), in particular in genes affecting the competitive ability of P. inhibens, including genes related to the production of the antibiotic tropodithietic acid (TDA) and bacterial cell-cell communication (e.g., quorum sensing). This work is significant as it reveals how the biofilm lifestyle might shape genome evolution in a cosmopolitan bacterium.

Transporter characterisation reveals aminoethylphosphonate mineralisation as a key step in the marine phosphorus redox cycle.

The planktonic synthesis of reduced organophosphorus molecules, such as alkylphosphonates and aminophosphonates, represents one half of a vast global oceanic phosphorus redox cycle. Whilst alkylphosphonates tend to accumulate in recalcitrant dissolved organic matter, aminophosphonates do not. Here, we identify three bacterial 2-aminoethylphosphonate (2AEP) transporters, named AepXVW, AepP and AepSTU, whose synthesis is independent of phosphate concentrations (phosphate-insensitive). AepXVW is found in diverse marine heterotrophs and is ubiquitously distributed in mesopelagic and epipelagic waters. Unlike the archetypal phosphonate binding protein, PhnD, AepX has high affinity and high specificity for 2AEP (Stappia stellulata AepX Kd 23 ± 4 nM; methylphosphonate Kd 3.4 ± 0.3 mM). In the global ocean, aepX is heavily transcribed (~100-fold>phnD) independently of phosphate and nitrogen concentrations. Collectively, our data identifies a mechanism responsible for a major oxidation process in the marine phosphorus redox cycle and suggests 2AEP may be an important source of regenerated phosphate and ammonium, which are required for oceanic primary production.

Pikeienuella piscinae gen. nov., sp. nov., a novel genus in the family Rhodobacteraceae.

A novel bacterium, designated strain RR4-56T, was isolated from a biofilter of a seawater recirculating aquaculture system. The 16S rRNA gene sequence analysis showed that the isolate was closely related to Halovulum dunhuangense YYQ-30T (92.6%), Albimonas donghaensis DS2T (91.3%), Pontivivens insulae GYSW-23T (91.3%), and Monaibacterium marinum C7T (90.9%), belonging to the family Rhodobacteraceae. The strain was aerobic, Gram-negative, rod-shaped, oxidase-positive, and catalase-negative. Its optimum temperature, pH, and salinity for growth were 25-30°C, pH 8.5, and 2-3% NaCl (w/v), respectively. Its growth occurred at 15-35°C, pH 5.0-9.5, and 0-7% NaCl (w/v). It contained ubiquinone-10 (Q-10), a respiratory quinone, and the major cellular fatty acids were 11-methyl C18:1 ω7c (31.9%), C18:1 ω6c (30.4%), and C19:0 cyclo ω8c (16.1%). The polar lipids present in the strain were phosphatidylglycerol, an unidentified phospholipid, and an unidentified aminolipid. The strain had one 4,373,045 bp circular chromosome with G + C contents of 65.9 mol% including 4,169 genes, 4,118 coding sequences (CDSs), 3 rRNAs, and 45 tRNAs. Genome annotation predicted some gene clusters related to the degradation of several types of organic matter such as protocatechuate, catechol, and phthalate. Based on the polyphasic characteristics, RR4-56T represents a novel genus and species in the family Rhodobacteraceae, for which the name Pikeienuella piscinae gen. nov., sp. nov. was proposed. The type strain is RR4-56T (= KCTC 52648T = DSM 107918T).

Marivivens aquimaris sp. nov., isolated from seawater.

A Gram-stain-negative, strictly aerobic, non-flagellated, rod-shaped bacterium, designated GSB7T, was isolated from seawater collected at the Yellow Sea coast of South Korea. Catalase and oxidase activities were positive. Growth occurred at pH 6.0-9.0 (optimum pH 7.0), 10-40 °C (optimum 30 °C) and with 0-8% NaCl (optimum 1-2%). Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain GSB7T belonged to the genus Marivivens, showing the sequence similarities of 96.3, 96.1, and 96.0% with Marivivens niveibacter HSLHS2T, Limimaricola hongkongensis DSM17492T, and Marivivens donghaensis AM-4T, respectively. The respiratory quinone was ubiquinone-10 and the major fatty acids were summed feature 8 (C18:1 ω7c and/or C18:1 ω6c), C18:1 ω7c 11-methyl, C16:0 and C10:0 3-OH. The polar lipids comprised phosphatidylglycerol, diphosphatidylglycerol, one unidentified aminolipid, and five unidentified lipids. The DNA G + C content calculated from the whole-genome sequence was 60.6 mol%. On the basis of phenotypic, chemotaxonomic and genotypic characteristics presented in this study, strain GSB7T is suggested to represent a novel species of the genus Marivivens, for which the name Marivivens aquimaris sp. nov. is proposed. The type strain is GSB7T (= KCTC 82026T = JCM 34042T).

Metabolic potential of the moderate halophile Yangia sp. ND199 for co-production of polyhydroxyalkanoates and exopolysaccharides.

Yangia sp. ND199 is a moderately halophilic bacterium isolated from mangrove samples in Northern Vietnam, which was earlier reported to grow on several sugars and glycerol to accumulate poly(hydroxyalkanoates) (PHA). In this study, the potential of the bacterium for co-production of exopolysaccharides (EPS) and PHA was investigated. Genome sequence analysis of the closely related Yangia sp. CCB-M3 isolated from mangroves in Malaysia revealed genes encoding enzymes participating in different EPS biosynthetic pathways. The effects of various cultivation parameters on the production of EPS and PHA were studied. The highest level of EPS (288 mg/L) was achieved using sucrose and yeast extract with 5% NaCl and 120 mM phosphate salts but with modest PHA accumulation (32% of cell dry weight, 1.3 g/L). Growth on fructose yielded the highest PHA concentration (85% of CDW, 3.3 g/L) at 90 mM phosphate and higher molecular weight EPS at 251 mg/L yield at 120 mM phosphate concentration. Analysis of EPS showed a predominance of glucose, followed by fructose and galactose, and minor amounts of acidic sugars.

Enhanced 2-keto-L-gulonic acid production by a mixed culture of Ketogulonicigenium vulgare and Bacillus megaterium using three-stage temperature control strategy.

As a key precursor of vitamin C, 2-keto-L-gulonic acid (2-KLG) was mainly produced from L-sorbose by mixed fermentation of Ketogulonicigenium vulgare and a helper strain (Bacillus spp.) with a low conversion rate for decades. The aim of this study was to enhance the 2-KLG production by co-culturing K. vulgare and Bacillus megaterium using three-stage temperature control (TSTC) strategy. By investigating the temperature effect on the 2-KLG fermentation, the optimum temperatures for the growths of K. vulgare and B. megaterium were 32 °C and 29 °C, respectively, while the optimum temperature for 2-KLG production was 35 °C. We developed a TSTC process: the temperature was kept at 32 °C during the first 16 h of fermentation, then decreased to 29 °C for the following 14 h, and maintained at 35 °C to the end of fermentation. By using this new process, the productivity and yield of 2-KLG from L-sorbose were obtained at 2.19 ± 0.19 g/L/h and 92.91 ± 1.02 g/L in 20-L fermentors for 5 batches, respectively, which were 22.35% and 6.02% higher than that of the control treatment (the single temperature of 29 °C). The increased cell density of K. vulgare during the exponential phase and the enhanced SDH activity (increased by 25.18% at 36 h, 17.14% at 44 h) in the production stage might be the reasons for enhanced 2-KLG conversion rate and yield. Our results demonstrated the feasibility of the TSTC strategy for 2-KLG production.

Peptide-Mediated Gene Transfer into Marine Purple Photosynthetic Bacteria.

Use of photosynthetic organisms is one of the sustainable ways to produce high-value products. Marine purple photosynthetic bacteria are one of the research focuses as microbial production hosts. Genetic transformation is indispensable as a biotechnology technique. However, only conjugation has been determined to be an applicable method for the transformation of marine purple photosynthetic bacteria so far. In this study, for the first time, a dual peptide-based transformation method combining cell penetrating peptide (CPP), cationic peptide and Tat-derived peptide (dTat-Sar-EED) (containing D-amino acids of Tat and endosomal escape domain (EED) connected by sarcosine linkers) successfully delivered plasmid DNA into Rhodovulum sulfidophilum, a marine purple photosynthetic bacterium. The plasmid delivery efficiency was greatly improved by dTat-Sar-EED. The concentrations of dTat-Sar-EED, cell growth stage and recovery duration affected the efficiency of plasmid DNA delivery. The delivery was inhibited at 4 °C and by A22, which is an inhibitor of the actin homolog MreB. This suggests that the plasmid DNA delivery occurred via MreB-mediated energy dependent process. Additionally, this peptide-mediated delivery method was also applicable for E. coli cells. Thus, a wide range of bacteria could be genetically transformed by using this novel peptide-based transformation method.